This invention relates to the in vitro production of human tumor angiogenesis factor from the human Burkitt lymphoma cell line Raji.
It has been known for some time that unless a solid tumor is provided with blood vessels by its host it remains small and dormant. The substance that is released by tumors and provides vascularization has been named tumor angiogenesis factor (TAF) by Dr. Judah Folkman of the Harvard Medical School. J. Exptl. Med. 133, 275-88 (1971); Cancer Res. 34, 2109-13 (1974).
Ever since it was recognized that tumor induced neovascularization represents the breaching of an important barrier in the control of tumor growth, considerable effort has been spent in searching for ways to inhibit neovascularization. It was early suggested by Dr. Folkman that blockade of this factor (inhibition of angiogenesis) might arrest solid tumors at a tiny diameter of a few millimeters (avascular phase). New Engl. J. Med. 285, 1182 (1971); J. Exptl. Med. 133, 275-88 (1971). One suggested approach was the raising of antibodies against TAF extracts for the production of an anti-serum. Folkman, Ann. Surg. 175 409-16 (1972); Phillips et al., Int. J. Cancer 17, 549-58 (1976). Another approach to inhibiting neovascularization which has received much attention recently consists in treatment of tumor cells with extracts of cartilage. Eisenstein et al, Am. J. Pathol. 73, 765-74 (1973); Sorgente et al., Lab. Invest. 32, 217-22 (1975); Eisenstein et al., Amer. J. Pathol. 81, 337-48 (1975); "Medical News", JAMA 232 (1), 14-15 (1975); Brem and Folkman, J. Exptl. Med. 141, 427-39 (1975); and Kuettner et al., U.S. Pat. No. 4,042,457. These efforts are limited, of course, to the amount of TAF which is available and suitable for test purposes.
TAF finds further use in the development of tests such as an angiogenic assay or a diagnostic screening test for neoplasia. Klagsbrun et al., Cancer Res. 36, 110-14 (1976); and Brem et al., Science 195, 880-81 (1977); Cancer 41, 239-44 (1978).
TAF also has been proposed as useful for wound healing, Rettusa et al, FASEB, Abstract No. 4309, 61st Ann. Meet., Apr. 1-8, 1977, Chicago, Ill.
Various human tumor cells have been reported heretofore as capable of elaborating TAF when measured by certain assays. Thus, extracts from human neoroblastoma, Wilms' tumor and human hepatoblastoma were found to contain TAF which was able to cause the formation of new blood vessels in the subcutaneous fascia of rats. Folkman et al., J. Exptl. Med. 133, 275-88 (1971).
So also, WI-38 embryonic lung, SV W126 (SV 40 virus transformed W126), glioblastoma, meningioma and HeLa cells (the latter only in suspension culture and not in monolayers) grown in T-75 tissue culture flasks were described as giving a positive TAF response as measured by bioassay on the chick chorioallantoic membrane (CAM). Folkman and Klagsbrun, Chapter 31 of "Fundamental Aspects of Neoplasia," at page 402, (Gottlieb et al, eds.), Springer-Verlag, New York, 1975.
Subsequently, extracts from hypernephroma, haemangioma and human kidney were similarly described as eliciting a TAF response which resulted in the growth of new capillaries in the subcutaneous fascia of rats. Phillips et al., Int. J. Cancer 17, 549-58 (1976).
In still other experiments, Hubler and Wolf demonstrated TAF responses from human cutaneous melanoma implanted in transparent hamster-cheek-pouch membranes. Cancer 38, 187-92 (1976).
All of the foregoing tumor cells are derived from fresh tumors or primary cultures except the WI-38, SV W126 and HeLa cells. Fresh tumors and primary cultures are not, however, generally suitable sources of TAF except for limited research purposes or small scale production. In order to provide a commercially significant source of TAF in terms of ready availability and adequate supply, production from a suitable established cell line is deemed necessary. As a practical matter, the cell line should have not only specific TAF activity, but it should also have good cell growth characteristics in terms of rapid growth, good suspension growth, adaptability to large-scale culture and economical nutrient requirements.
The terms "cell line" and "established cell line" are used herein in conformance with the definitions published by Federoff in the Tissue Culture Association Manual, Vol. 1, No. 1, pp. 53-57 (1975).